Dataset Information

Anti-HER2 monoclonal antibodies intensify the susceptibility of human gastric cancer cells to etoposide by promoting apoptosis, but not autophagy.

ABSTRACT:

Background

Gastric cancer (GC) is a multifactorial disease with high mortality. Anti-HER2 therapy is a promising strategy in GC treatment and trastuzumab was approved by FDA (Food and Drug Administration) as the first and the second line of treatment of the disease.

Purpose

The aim of the study was to examine the effectiveness of a combination of etoposide with trastuzumab or pertuzumab in AGS gastric cancer cells and breast cancer cells such as MCF-7, MDA-MB-231 and HCC1954.

Methods and findings

The cytotoxic effects of the tested compounds against gastric and breast cancer cells were checked by MTT (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide) assay. The anti-proliferative potential was analyzed by the incorporation of [3H]-thymidine into DNA. Fluorescent microscopy and flow cytometry was used to demonstrate the effect of the compounds on apoptosis. The mitochondrial membrane potential, and the activity of caspase-8 and caspase-9 were assessed. Autophagosomes and autolysosomes formation was checked by flow cytometry. The concentrations of Beclin-1, LC3A and LC3B were performed using ELISA. The expression of LC3A/B was also determined. The results from our study proved that the combination of etoposide with anti-HER2 antibodies was not cytotoxic against breast cancer cells, whereas the combination of etoposide with anti-HER2 antibodies decreased viability and DNA biosynthesis in gastric cancer cells. The interaction of etoposide with pertuzumab or trastuzumab induced programmed cell death via extrinsic and intrinsic apoptotic pathways in AGS gastric cancer cells, but did not affect autophagy, where a decrease of Beclin-1, LC3A and LC3B was observed in comparison with the untreated control.

Conclusions

The study demonstrated that etoposide (12.5 μM) with pertuzumab represent a promising strategy in gastric cancer treatment, but further in vivo examinations are also required.

SUBMITTER: Gornowicz A

PROVIDER: S-EPMC8389407 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Publications

Anti-HER2 monoclonal antibodies intensify the susceptibility of human gastric cancer cells to etoposide by promoting apoptosis, but not autophagy.

Gornowicz Agnieszka A Szymanowski Wojciech W Czarnomysy Robert R Bielawski Krzysztof K Bielawska Anna A

PloS one 20210826 8

<h4>Background</h4>Gastric cancer (GC) is a multifactorial disease with high mortality. Anti-HER2 therapy is a promising strategy in GC treatment and trastuzumab was approved by FDA (Food and Drug Administration) as the first and the second line of treatment of the disease.<h4>Purpose</h4>The aim of the study was to examine the effectiveness of a combination of etoposide with trastuzumab or pertuzumab in AGS gastric cancer cells and breast cancer cells such as MCF-7, MDA-MB-231 and HCC1954.<h4>M ...[more]

PMID: 34437575

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Project description:BackgroundOne-third of breast cancers display amplifications of the ERBB2 gene encoding the HER2 kinase receptor. Trastuzumab, a humanized antibody directed against an epitope on subdomain IV of the extracellular domain of HER2 is used for therapy of HER2-overexpressing mammary tumors. However, many tumors are either natively resistant or acquire resistance against Trastuzumab. Antibodies directed to different epitopes on the extracellular domain of HER2 are promising candidates for replacement or combinatorial therapy. For example, Pertuzumab that binds to subdomain II of HER2 extracellular domain and inhibits receptor dimerization is under clinical trial. Alternative antibodies directed to novel HER2 epitopes may serve as additional tools for breast cancer therapy. Our aim was to generate novel anti-HER2 monoclonal antibodies inhibiting the growth of breast cancer cells, either alone or in combination with tumor necrosis factor-? (TNF-?).MethodsMice were immunized against SK-BR-3 cells and recombinant HER2 extracellular domain protein to produce monoclonal antibodies. Anti-HER2 antibodies were characterized with breast cancer cell lines using immunofluorescence, flow cytometry, immunoprecipitation, western blot techniques. Antibody epitopes were localized using plasmids encoding recombinant HER2 protein variants. Antibodies, either alone or in combination with TNF-?, were tested for their effects on breast cancer cell proliferation.ResultsWe produced five new anti-HER2 monoclonal antibodies, all directed against conformational epitope or epitopes restricted to the native form of the extracellular domain. When tested alone, some antibodies inhibited modestly but significantly the growth of SK-BR-3, BT-474 and MDA-MB-361 cells displaying ERBB2 amplification. They had no detectable effect on MCF-7 and T47D cells lacking ERBB2 amplification. When tested in combination with TNF-?, antibodies acted synergistically on SK-BR-3 cells, but antagonistically on BT-474 cells. A representative anti-HER2 antibody inhibited Akt and ERK1/2 phosphorylation leading to cyclin D1 accumulation and growth arrest in SK-BR-3 cells, independently from TNF-?.ConclusionsNovel antibodies against extracellular domain of HER2 may serve as potent anti-cancer bioactive molecules. Cell-dependent synergy and antagonism between anti-HER2 antibodies and TNF-? provide evidence for a complex interplay between HER2 and TNF-? signaling pathways. Such complexity may drastically affect the outcome of HER2-directed therapeutic interventions.

Dataset Information

Anti-HER2 monoclonal antibodies intensify the susceptibility of human gastric cancer cells to etoposide by promoting apoptosis, but not autophagy.

Background

Purpose

Methods and findings

Conclusions

Publications

Anti-HER2 monoclonal antibodies intensify the susceptibility of human gastric cancer cells to etoposide by promoting apoptosis, but not autophagy.

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